The present invention relates to the coupling of targeting molecules to lipid vesicles. More particularly, methods of coupling targeting molecules, e.g., proteins, to the lipid bilayers of lipid vesicles or liposomes are disclosed which allow a substantial number of targeting molecules to be coupled to each vesicle without decreasing the stability of the vesicle bilayers. Products made using these methods, including targeted vesicles, are also disclosed.
Almost since their discovery, the possible use of lipid vesicles, primarily the phospholipid lipid vesicles commonly known as liposomes, as agents for carrying materials to specific cell groups or organs in the body has been discussed. In order to accomplish this, targeting molecules must be attached to the surface of the vesicle. These targeting molecules, when reacted with, or embedded in, the outer surface of the lipid vesicles, cause a reaction between the targeting molecules and a desired feature of the targeted cell group, e.g., a cell surface receptor or an antigenic feature on a cell. Currently used methods of coupling phospholipid vesicles to proteins rely primarily on reacting a bifunctional reagent with the amino group of phosphatidylethanolamine included in the bilayer and coupling the protein to the other end of the bifunctional molecule in the lipid layer. However, this procedure leads to many problems including breakdown of the lipid layer and resulting instability of the lipid vesicles. Another approach has been to couple targeting proteins to palmitic acid chains in the bilayer. However, only a few targeting molecules can be introduced in this way and the same complications arise. Accordingly, only small quantities of targeted molecules have been able to be attached to the vesicle without vesicle breakdown.
Similar coupling procedures can be used for in vitro diagnostic agents, particularly immunodiagnostics, as are used for targeting vesicles. As expected, the same type of problems have been observed concerning vesicle stability.
Accordingly, an object of the invention is to provide a method of coupling proteins or other targeting molecules to lipid vesicles which providing high levels of coupling while reducing vesicle breakdown.
A further object of the invention is to provide a method of coupling targeting molecules to lipid vesicles which can be used with both phospholipid and nonphospholipid vesicles.
Another object of the invention is to provide a method of coupling proteins to lipid vesicles which does not cause a loss of activity of the protein function.
These and other objects and features of the invention will be apparent from the following description and the claims.